Plant Science for Global Challenges

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Month: September 2016

The 1000 plants initiative (1KP) is a multidisciplinary consortium aiming to generate large-scale gene sequencing data for over 1000 species of plants. Included in these species are those of interest to agriculture and medicines, as well as green algae, extremophytes and non-flowering plants. The project is funded by several supporters, and has already generated many published papers.

DS: This has been an unparalleled opportunity to reveal and understand the genes that have led to the plant diversity we see around us. We were able to study plants that were pivotal in terms of plant evolution but which have not previously been included in sequencing projects as they are not considered important economically

GW: The project was funded by the Government of Alberta and the investment firm Musea Ventures to raise the profile of the University of Alberta. Notably there was no requirement by the funders to sequence any particular species. I was able to ask the plant science community what the best possible use of these resources would be. The community was in full agreement that the money should be used to sample plant diversity.

Hopefully our work will change the thinking at the funding agencies regarding the value of sequencing biodiversity.

What techniques were utilized in this project to carry out the research?

GW: Complete genomes were too expensive to sequence. Many plants have unusually large genomes and de novo assembly of a polyploid genome remains difficult. To overcome this problem, we sequenced transcriptomes. However, this made our sample collection more difficult as the tissue had to be fresh. In addition, when we started the project, the software to assemble de novo transcriptomes did not work particularly well. I simply made a bet that these problems would be solved by the time we collected the samples and extracted the RNA. For the most part that’s what happened, although we did end up developing our own assembly software as well!

The 1KP initiative is an international consortium. How has the group evolved over time and what benefits have you seen from having this diverse set of skills?

GW: 1KP would not be where it is today without the participation of scientists around the world from many different backgrounds. For example, plant systematists who defined species of interest and provided the tissue samples worked alongside bioinformaticians who analyzed the data, and gene family experts who are now publishing fascinating stories about particular genes.

DS: One of the great things about this project is how it has evolved over time as new researchers became involved. There is no restriction on who can take part, which species can be studied or which questions can be asked of the data. This makes the 1KP initiative unique compared to more traditionally funded projects.

GW: We continually encouraged others to get involved and mine our data for interesting information. We did a lot of this through word of mouth and ended up with some highly interesting, unexpected discoveries. For example, an optogenetics group at MIT and Harvard used our data to develop new tools for mammalian neurosciences. This really highlights the importance of not restricting the species we study to those of known economic importance.

According to ISI outputs from this research, two of the most highly cited papers from 1KP are here and here.

You aimed to investigate a highly diverse array of plants. How many plants of the major phylogenetic groups have now been sequenced, and are you still working on expanding the data set?

DS: A lot of thought went into the species selection. We aimed for proportional representation (by number of species) of the major plant groups. We also aimed to represent the morphological diversity of those groups.

Has this project lead to any breakthroughs in our understanding of the phylogeny of plants?

DS: This will be the first broad look at what the nuclear genome has to tell us, and the first meaningful comparison of large nuclear and plastid data sets. However, due to rapid evolution plus extinction, many parts of the plant evolutionary tree remain extremely difficult to solve.

One significant breakthrough was the discovery of horizontal gene transfer from a hornwort to a group of ferns. This was unexpected and very interesting in terms of the ability of those ferns to be able to accommodate understory habitats.

GW: With regard to horizontal gene transfer, there are papers in the pipeline that will illustrate the discovery of even more of these events in other species. We have also studied gene duplications at the whole genome and gene family level. This is the most comprehensive survey ever undertaken, and people will be surprised at the scale of the discoveries. However, we will be releasing our findings shortly as part of a series and it would be unwise for us to give the story away here! Keep a look out for these!

This week’s post was written by Katie Tomlinson, a PhD student at the University of Bristol, UK, who spent three months as an intern at the National Crops Resource Research Institute in Uganda. She fills us in on the important research underway at the Institute, and how they communicate their important results to local farmers and benefit rural communities.

Over the summer, I had a great time at the National Crops Resources Research Institute (NaCRRI) in Uganda. I’m currently in the second year of my PhD at the University of Bristol, UK, where I’m researching how the cassava brown streak disease (CBSD) viruses are able to cause symptoms, replicate and move inside plants. I was lucky enough to be given a placement at NaCRRI as part of the South West Doctoral Training Partnership Professional Internship for PhD Students (PIPS) scheme, to experience the problem for myself, see the disease in the field, meet the farmers affected and investigate the possible solutions.

Cassava is a staple food crop for approximately 300 million people in Africa. It is resilient to seasonal drought, can be grown on poor soils and harvested when needed. However, cassava production is seriously threatened by CBSD, which causes yellow patches (chlorosis) to form on leaves and areas of tubers to die (necrosis), rot and become inedible.

Despite being identified in coastal Tanzania 80 years ago, CBSD has only been a serious problem for Uganda in the last 10 years, where it was the most important crop disease in 2014–2015. The disease has since spread across East Africa and threatens the food security of millions of people.

NaCRRI is a government institute, which carries out research to protect and improve the production of key crops, including cassava. The focus is on involving farmers in this process so that the best possible crop varieties and practices are available to them. Communication between researchers and farmers is therefore vital, and it was this that I wanted to assist with.

When I arrived I was welcomed warmly into the root crop team by the team leader Dr Titus Alicai, who came up with a whole series of activities to give me a real insight into CBSD. I was invited to field sites across Uganda, where I got to see CBSD symptoms in the flesh! I helped to collect data for the 5CP project, which is screening different cassava varieties from five East and Southern African countries for CBSD and cassava mosaic disease (CMD) resistance. I helped to score plants for symptoms and was fascinated by the variability of disease severity in different varieties. The main insight I gained is that the situation is both complex and dynamic, with some plants appearing to be disease-free while others were heavily infected. There are also different viral strains found across different areas, and viral populations are also continually adapting. The symptoms also depend on environmental conditions, which are unpredictable.

I also got to see super-abundant whiteflies, which transmit viruses, and understand how their populations are affected by environmental conditions. These vectors are also complex; they are expanding into new areas and responding to changing environmental conditions.

It has been fascinating to learn how NaCRRI is tackling the CBSD problem through screening different varieties in the 5CP project, breeding new varieties in the NEXTGEN cassava project, providing clean planting material and developing GM cassava.

And there’s the human element…

In each of these projects, communication with local farmers is crucial. I’ve had the opportunity to meet farmers directly affected, some of whom have all but given up on growing cassava.

Challenging communications

Communicating has not been easy, as there are over 40 local languages. I had to adapt and learn from those around me. For example, in the UK we have a habit of emailing everything, whereas in Uganda I had to talk to people to hear about what was going on. This is all part of the experience and something I’ll definitely be brining back to the UK! I’ve had some funny moments too… during harvesting the Ugandans couldn’t believe how weak I was; I couldn’t even cut one cassava open!

Real world reflections

I’m going to treasure my experiences at NaCRRI. The insights into CBSD are already helping me to plan experiments, with more real-world applications. I can now see how all the different elements (plant–virus–vector–environment–human) interact, which is something you can’t learn from reading papers alone!

Working with the NaCRRI team has given me the desire and confidence to collaborate with an international team. I’ve formed some very strong connections and hope to have discussions about CBSD with them throughout my PhD and beyond. It’s really helped to strengthen collaborations between our lab work in Bristol and researchers working in the field on the disease frontline. This will help our research to be relevant to the current situation and what is happening in the field.

Science is not a solo endeavour but a social one, and the most social part is conference attendance. Regardless of their other strengths and weaknesses, scientific meetings are critical for encouraging researchers early in their careers.

Unquestionably, one of the most enjoyable aspects of being a journal editor is the opportunity to attend conferences. While the average scientist may get to one or two scientific meetings a year, we try to get to many more — and so are in a good position to compare the different styles of meeting, and to try to understand what makes a conference not just good, but great.

Mainly, it is the people who are attending. Meetings are exactly what the name implies: an opportunity to meet colleagues and discuss science. But there are many factors that determine who will attend a conference, and whether they will get to talk constructively while they are there. Location is important. Many scientific conferences are held in places well worth visiting in their own right. Last year’s International Plant Molecular Biology Congress, for example, was held near Iguazú Falls, Brazil; the XIV Cell Wall Meeting was held this year on the Greek island of Crete; and, next year, the Plant Biology 2017 conference of the American Society of Plant Biology (ASPB) will be in Honolulu, Hawaii. However, as much as exotic locations may be a draw for participants, the long and expensive journeys can be a deterrent.

An additional factor is the breadth, or narrowness, of focus of a meeting, which affects both its size and atmosphere. Larger meetings with a broad range of topics guarantee that there will be something of interest to everyone. These can be superb at giving a broad view of the important questions currently being addressed in a field, and usually have presentations by impressive well-known and well-practiced speakers. However, it can be difficult to meet all the people with whom you want to chat without considerable dedication and forward planning.

You often see a reluctance in speakers to present new and unpublished work at larger meetings. For that, smaller meetings come into their own, where a more tightly defined community makes it more appealing to share confidences in a room perceived to be full of ‘friends’. If the location is remote, so much the better, as it forces that community closer together. The summertime masters of such meetings are the Gordon Research Conferences, which are often (though not exclusively) held in out-of-season New England boarding schools — two of which, this year, are the Plant Molecular Biology and Plant & Microbial Cytoskeleton meetings. In the winter, there are the Keystone Symposia, which have the added attraction of afternoons left free for skiing. In fact, the conversations had while trapped on a ski lift can often be the most scientifically productive of the whole event.

More focused meetings will usually give attendees the opportunity to attend every talk, but larger conferences frequently host parallel sessions to allow many more topics to be discussed. Successfully presenting parallel sessions is hard. Ideally the topics covered should overlap so little that every attendee would wish to attend one session, and one session only — a goal never fully achieved, and rarely even approached. Instead, attendees must pick the talks that they most want to see, which are often presented in different sessions, leading to a lot of distracting crowd movement between talks. For sessions to remain synchronized, speakers must keep strictly to their allotted time — again something so difficult to achieve that it rarely, if ever, happens.

At its heart, the main point of a scientific conference is not to visit interesting places, to catch up with old friends, to party with colleagues (although much partying does occur), or even to listen to high-profile scientists lecture on their work. All these are important aspects of a successful conference, but its central function is to bring people together to discuss their own studies. Where this happens most is at the poster sessions — the great equalizer of any scientific conference..

GPC New Media Fellow Sarah Jose presents a poster at a conference

However lofty the professor or junior the student, with a poster everyone can present their work on an equal level, open to the criticism of all. They are the soul of any good conference, but they are the most difficult aspect to organize successfully. Ideally the posters should all be in one place rather than spread out over a number of rooms, to avoid some groups getting ignored. The posters need to be arranged close enough together that when the session is in full swing there is a throng and hubbub of chatter, but not so closely packed that posters are blocked by people reading the next one over. It is also vital that there is enough space to move freely between posters without having to squeeze past huddles of scientists talking with the presenters. Above all, posters must be available for long enough that conference-goers can read all that are relevant to them. Therefore poster rooms need to be open throughout the conference, not just during designated sessions, and all posters should be available for the whole conference, not taken down halfway through to make way for a second batch.

Posters provide some of the first opportunities that early-career scientists have to present their research. It is therefore always good to see conferences enhancing their status in some way. The simplest is the awarding of prizes for the ‘best’ posters, judged as much for the clarity of presentation as for the story being told. Some conferences have started to schedule ‘flash talks’, selecting presenters to give a short description of their work, and serving as an advert for their posters. This commonly takes the format of five-minute presentations with no more than three slides — but ‘slam’ sessions are also possible, where a single minute is allocated to each speaker. A variation of this occurred at the recent ASPB Plant Biology 2016 meeting in Austin, Texas: early-stage researchers were helped to video ‘elevator pitches’ about their work, which can now be seen on the Plantae YouTube channel. It is also encouraging to see that the New Phytologist Trust will again be holding a Next Generation Scientists symposium next year, following on from the successful inaugural meeting in 2014.

The planning, organization and execution of a scientific meeting requires as much skill, enthusiasm and innovation as any other part of the scientific endeavour. After all, a good conference brings scientists together to discuss ideas, initiate collaborations and forge friendships that can last for entire careers, and sometimes longer.

This week we spoke to Dr. Winfried S. Peters from Indiana University/Purdue University Fort Wayne (IPFW). His research mainly focuses on the biomechanics of plant cells, which led him to take a second look at some of the ideas of botanists in the 19th and early 20th century and use modern techniques to make exciting new discoveries.

Could you begin by describing your research interests?
I am interested in the biophysical aspects of the physiology of plants and animals. In plants, my research focuses on the mechanics of growth and morphogenesis, and on the cell biology of long-distance transport in the phloem. For both topics, a solid background in the history of the field can be quite helpful – I love studying the old literature to reconstruct the ideas botanists had a century or two ago regarding the functioning of plants.

At the recent New Phytologist Symposium, entitled “Colonization of the terrestrial environment 2016”, you presented fascinating work on the sieve tubes of kelp, which resemble the phloem tubes of vascular plants. What is the purpose of these tubes?
In large photosynthetic organisms, not all parts of the body are truly autototrophic. Some tissues produce more material by photosynthesis than they need, while others produce less than they require or none at all– think of green leaves and growing root tips. Over-producing tissues can act as sources and export photoassimilates to needy sink tissues. Sieve tubes are arrays of tubular cells that mediate this exchange, enabling the rapid movement of photosynthate-rich cytoplasm between sources and sinks.

What techniques did you utilize to investigate the function of these tubes, and what did this reveal?
During my recent sabbatical, I became involved in this project in the lab of my friend and long-term collaborator, Professor Michael Knoblauch. Michael heads the Franceschi Microscopy and Imaging Center at Washington State University, where we studied sieve tubes of the Bull Kelp (Nereocystis luetkeana) using a variety of state-of-the-art microscopy techniques. Most importantly, we employed fluorescent dyes to visualize transport in sieve tube networks. To do this, one needs to work with intact kelp, which is demanding given a thallus size of 12 meters and more. So we moved to Bamfield Marine Sciences Centre on Vancouver Island, where Bull Kelp is a ‘common weed’.

A particularly important result was the pressure-induced reversal of the flow direction in sieve tubes and across sieve plates. This was in line with Ernst Münch’s (1876-1946) theory, who suggested that sieve tube transport was driven by osmotically generated pressure gradients.

An intact Nereocystis luetkeana is kept in a tank (right) while sieve tube transport is studied using a fluorescence microscope. Photo credit: Michael Knoblauch.

How do the biomechanics of the kelp sieve tubes differ from the phloem tubes of higher plants?Regarding cytoplasmic translocation, there doesn’t seem to be a difference – in higher plants as in kelps, the contents of the sieve tubes move in bulk flow – but wounding responses differ drastically. After wounding, we found that kelps have a massive swelling of the walls, which reduced the sieve tube diameter by more than 70%. By injecting silicon oil into severed kelp sieve tubes we demonstrated that wall swelling was fully reversible, and that the swelling state of the walls depended on intracellular pressure.

Sieve wall tubes swell after wounding due to changes in intracellular pressure. (Images taken from video below).

Have reversible wall-swelling reactions been observed in other species, and what are the implications of this finding?
We have observed the wall-swelling response in all kelp species examined. Ironically, there is no shortage of drawings and photographs of kelp sieve tubes with swollen walls in the literature over the last 130 years; however, the dynamics of cell behavior remained hidden in plain sight because fixed tissue samples rather than fully functional, whole organisms were studied. Consequently, sieve tubes with swollen walls were misinterpreted as senescent cells. There also are publications on turgor-dependent cell wall swelling in red and green algae, but these ceased around 1930.

Afterwards, wall swelling was completely forgotten, judging from the textbooks. This is remarkable, as Wilhelm Hofmeister (1824-1877), often celebrated as a founding father of plant biomechanics, denied a significant role for osmotic processes in the generation of turgor, the hydrostatic pressure within plant cells. Rather, he maintained that living cells were pressurized by the swelling of their walls. The example of the kelp sieve tube shows how easy it is to remain unaware of wall swelling when it happens right before our eyes. Maybe we should take Hofmeister’s idea seriously once again?

What are the evolutionary implications of your work?
Brown algae and vascular (land) plants are only remotely related, and their sieve tube networks certainly evolved independently of each other. It seems surprising that such sophisticated structures, which serve a complex function that integrates the physiology of the entire organism, have evolved at least twice, but think again. Real cells are not embedded in a totally homogeneous environment, and neither is the cytoplasm within the cell a homogeneous solution. Thus every cell experiences gradients of solute concentrations along its inner and/or outer surface. As a consequence, differential water fluxes across the plasma membrane will occur, resulting in movements of the cell contents. In other words, Münch flow, the cytoplasmic bulk flow driven by osmotically generated pressure gradients, is not a peculiar process operating specifically in sieve tubes, but a ubiquitous phenomenon. Sieve tubes consist of cells that simply do the things cells do, just a little more efficiently as usual. In this view, the repeated convergent evolution of sieve tube networks is not really unexpected.

But kelps resemble land plants in other ways too. As in land plants, kelp cell walls are made of cellulose (at least partly), kelp cells are connected through plasmodesmata, and the kelp life-cycle is a sporophyte-dominated alternation of generations. Evidently, none of these features represents a specific adaptation to life on dry land.

Wound responses including wall swelling in a sieve tube of Nereocystis luetkeana. (Watch for the rapid cell wall swelling between 11 and 14 seconds in!) This video was taken by Professor Michael Knoblauch in collaboration with Dr Winfried S. Peters.

If you’d like to know more about this fascinating work, it was been published in the following articles: